CSB349H1_08SPRING1_246 - Copy

CSB349H1_08SPRING1_246 - Copy - CSB349H1S Term Test 1...

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CSB349H1S Term Test 1 Answer Key 1 1. The human gene VT is expressed in the brain and heart during early development, but in zebrafish there are two copies of this gene called VT-A and VT-B. In zebrafish, the VT-A gene is expressed only in the brain while the VT-B gene is expressed only in the heart. (a) Briefly describe how the duplication-degeneration-complementation (DDC) model of genomic evolution can explain the diverged expression of the two VT genes in zebrafish compared to the human VT expression pattern. (3 marks) - Ancestral VT gene is duplicated in the teleost lineage after the divergence of mammalian (human) and teleost (zebrafish) lineages (1 mark) - After the genes have duplicated, there is a loss of sequence conservation in the two separate regulatory regions (mutations accumulated differently over evolutionary time) (1 mark) (“degenerate” = 0.5 marks) - Changes in regulatory sequences of the two genes leads to changes in gene expression (VT-A in brain and VT-B in heart), such that they compliment one another with regards to the ancestral gene expression pattern (could also say subfunctionalization) (1 mark) (b) What experiment in zebrafish embryos would you perform to prove that the upstream regulatory regions of VT-A and VT-B have evolved separate functions in controlling gene expression in zebrafish embryos in vivo? ( Be sure to describe the basic steps in the experiment and the predicted results ) (3 marks) - Engineer separate enhancer-reporter constructs using upstream regulatory regions from gene VT-A (0.5 marks) and gene VT-B (0.5 marks) as well as a reporter (e.g. EGFP) (0.5 marks) - Generate trangenic zebrafish embryos for each construct (0.5 mark) - Show that each enhancer can direct the expression of reporter to either brain (VT-A) or heart (VT-B) (0.5 marks each) (c) Using zebrafish transgenic technology, describe how you would prove that the expression pattern of the human VT gene is likely to be most similar to the ancestral vertebrate expression pattern before DDC has taken place in the fish lineage. ( Be sure to describe the basic steps in the experiment and the predicted results ) (3 marks) - Engineer enhancer-reporter construct (0.5 marks) using human VT gene (0.5 marks) upstream regulatory region and reporter (0.5 marks) - Generate zebrafish transgenics (0.5 marks) - Expect to see reporter expression in brain AND heart (1 mark) 2. Promoter clearance is defined by the disassembly of most of the components of the pre-initiation complex and the synthesis of a short stretch of ~30 nucleotides of new mRNA by RNA Pol II, after which transcription is stalled. (a) What are the 2 main factors that cause RNA Pol II stalling during this phase of transcription?
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  • Fall '10
  • V.Tropepe,A.Moses
  • DNA, transposase

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